Flow cytometers are powerful analytical tools that allow the multi-parametric analysis of up to thousands of particles (such as cells on a cell-by-cell basis) per second. In a flow cytometer, the particles, tagged with fluorescence markers, flow past laser beams in single file. Typically, the parameters that are analyzed include light scatter and fluorescence signals, generated by interaction of the particles with light sources in the flow cytometer. Fluorescence markers, or fluorochromes, may be inherent to the particle or may be added by a user (e.g., researchers or clinicians) to tag specific cellular structures in the sample such as nucleic acids or proteins, or to follow specific cellular processes such as cross-membrane calcium or pH fluxes.
The demand for multi-color and multi-parameter analysis, along with recent advances in optics, electronics, and signal processing, has driven the development of multi-laser, multi-detector systems that can measure up to 30 or more fluorescence signals simultaneously. To satisfy this demand, the typical approach of flow cytometer manufacturers has been to add lasers and detectors to the optical bench of the instrument in proportion to the number of parameters to be measured. This approach assumes that each additional parameter measured will be labeled with a particular fluorochrome requiring a unique detector for analysis. However, this approach greatly increases the size and complexity of the flow cytometer. Further, the price and difficulty of setting up, operating, and maintaining such a flow cytometer quickly moves the system out of reach for most cytometry users who are interested in performing their own analyses. Thus, there is a need in the flow cytometry analysis field to create an improved method and system for detecting fluorochromes in a flow cytometer. This invention provides such improved an improved method.